Link protein is a glycoprotein found in all hyaline cartilage examined to date. It interacts specifically with a decasaccharide segment of hyaluronic acid and stabilizes the binding of proteoglycan monomers to hyaluronic acid to form aggregates and appears to interact with both proteoglycan and hyaluronic acid (Hardingham 1979; Franzen et al, 1981; Day et al, 1999). Mice lacking link protein develop dwarfism and craniofacial abnormalities, most mice die shortly after birth, changes of gonadal tissues are not recorded (Watanabe and Yamada 1999).
General function
Comment
Binette et al. (1994) showed that Link protein is ubiquitously expressed in non-cartilaginous tissues where it enhances and stabilizes the interaction of proteoglycans with hyaluronic acid.
Periovulatory expression of hyaluronan and proteoglycan link protein 1 (Hapln1) in the rat ovary: hormonal regulation and potential function. Liu J et al. Periovulatory follicular matrix plays an important role in cumulus-oocyte complex (COC) expansion, ovulation, and luteal formation. Hyaluronan and proteoglycan link protein 1 (HAPLN1), a component of follicular matrix, was shown to enhance COC expansion in vitro. However, the regulatory mechanisms of periovulatory expression of Hapln1 and its role in periovulatory granulosa cells have not been elucidated. We first determined the periovulatory expression pattern of Hapln1 using pregnant mare serum gonadotropin/human chorionic gonadotropin (PMSG/hCG)-primed immature rat ovaries. Hapln1 expression was transiently induced both in intact ovaries and granulosa cells at 8 h and 12 h after hCG injection. This in vivo expression of Hapln1 was recapitulated by culturing preovulatory granulosa cells with hCG. The stimulatory effect of hCG was blocked by inhibition of protein kinase A, phosphatidylinositol-dependent kinase, p38 MAPK, epidermal growth factor signaling, and prostaglandin synthesis, revealing key mediators involved in LH-induced Hapln1 expression. In addition, knockdown of Runx1 and Runx2 expression by small interfering RNA or inhibition of RUNX activities by dominant-negative RUNX decreased hCG or agonist-induced Hapln1 expression. Chromatin immunoprecipitation assays verified the in vivo binding of RUNX1 and RUNX2 to the Hapln1 promoter in periovulatory granulosa cells. Luciferase reporter assays revealed that mutation of the RUNX binding sites completely obliterated the agonist-induced activity of the Hapln1 promoter. These data conclusively identified RUNX proteins as the crucial transcription regulators for LH-induced Hapln1 expression. Functionally, treatment with HAPLN1 increased the viability of cultured granulosa cells and decreased the number of the cells undergoing apoptosis, whereas knockdown of Hapln1 expression decreased granulosa cells viability. This novel finding indicates that HAPLN1 may promote periovulatory granulosa cell survival, which would facilitate their differentiation into luteal cells.
Link protein is important for cumulus expansion possibly by stabilizing an aggregate structure of the hyaluronic acid-rich matrix (Kobayashi et al, 1999).Camaioni et al, (1996) propose that the dermatan sulfate proteoglycan and a approximately 46-kDa protein (same size as link-protein) synthesized by the cumulus cells form similar ternary complexes that are necessary for retaining HA in the COC matrix and hence are required for successful COC expansion.
Expression regulated by
FSH, LH, Growth Factors/ cytokines
Comment
Addition of FSH to the media of cumulus cells in culture resulted in increased rate of link protein synthesis (Kobayashi et al, 1999).And Sun GW, et al 2003 reported that Follicle-Stimulating Hormone and Insulin-Like Growth Factor I Synergistically Induce Up-Regulation of Cartilage Link Protein (Crtl1) via Activation of Phosphatidylinositol-Dependent Kinase/Akt in Rat Granulosa Cells.
Ovarian localization
Oocyte, Cumulus
Comment
Link protein staining was found in most of the oocytes but not in granulosa cells of immature rat follicles before PMSG treatment. After stimulation with PMSG and HCG link protein was found in the cytoplasm and extracellular matrix of cumulus cells. Furthermore, cumulus cells deposit link protein into the extracellular matrix in vitro (Kobayashi et al, 1999).
Follicle stages
Preovulatory
Comment
Immunohistochemical analysis showed the extracellular matrix of expanded cumulus oocyte complexes markedly stained for link protein, cumulus cells from immature follicles were not stained (Kobayashi et al, 1999).